Optimizing the mass production of Clonostachys rosea by liquid-state fermentation

Abstract

Optimization of the culture medium and the entire mass production process for various fungi in liquid medium has been studied. However, the technology is not well developed for Clonostachys rosea, a biocontrol agent against various plant pathogens due to its antagonistic capacity to act as a hyperparasite, compete for nutrients and space, and induce plant resistance to pathogens. In this study, we aimed to optimize the culture medium and to standardize parameters that may interfere with the production of C. rosea conidia in a liquid-state fermentation system. Culturing was performed in 250-mL Erlenmeyer flasks shaken for 7 days, followed by planned experimental methodology to reduce the number of analyses and consumable costs. Benchtop bioreactor tests with the optimized medium were performed. Glucose and sucrose were evaluated as carbon sources. Initially, the effects of temperature, pH, photoperiod, carbon:nitrogen ratio and water activity on inoculum production were evaluated, with the pH and photoperiod being factors that contributed to conidial production. Optimization of the fermentation conditions was performed using a central composite rotational design (CCD) with a wider range of pH values and photoperiods. The remaining variables were fixed according to the previous assay. Colony-forming unit (CFUs), biomass production and conidial viability were evaluated, and glucose was used as a carbon source to enhance conidial production. The optimized conditions that resulted in a maximum yield of conidia (1.78 × 107 conidia mL−1), dried biomass (0.558 g) and CFUs (5.15 × 106 CFUs mL−1) were a pH value of 4 and a photoperiod of 12 h.